title: BIM(EL), an intrinsically disordered protein, is degraded by 20S proteasomes in the absence of poly-ubiquitylation.
abstract: BIM-extra long (BIM(EL)), a pro-apoptotic BH3-only protein and part of the BCL-2 family, is degraded by the proteasome following activation of the ERK1/2 signalling pathway. Although studies have demonstrated poly-ubiquitylation of BIM(EL) in cells, the nature of the ubiquitin chain linkage has not been defined. Using ubiquitin-binding domains (UBDs) specific for defined ubiquitin chain linkages, we show that BIM(EL) undergoes K48-linked poly-ubiquitylation at either of two lysine residues. Surprisingly, BIM(EL)ΔKK, which lacks both lysine residues, was not poly-ubiquitylated but still underwent ERK1/2-driven, proteasome-dependent turnover. BIM has been proposed to be an intrinsically disordered protein (IDP) and some IDPs can be degraded by uncapped 20S proteasomes in the absence of poly-ubiquitylation. We show that BIM(EL) is degraded by isolated 20S proteasomes but that this is prevented when BIM(EL) is bound to its pro-survival target protein MCL-1. Furthermore, knockdown of the proteasome cap component Rpn2 does not prevent BIM(EL) turnover in cells, and inhibition of the E3 ubiquitin ligase β-TrCP, which catalyses poly-Ub of BIM(EL), causes Cdc25A accumulation but does not inhibit BIM(EL) turnover. These results provide new insights into the regulation of BIM(EL) by defining a novel ubiquitin-independent pathway for the proteasome-dependent destruction of this highly toxic protein.
authors.affiliation: Laboratory of Molecular Signalling, The Babraham Institute, Babraham Research Campus, Cambridge, CB22 3AT, UK.
authors.forename: Ceri M
authors.initials: CM
authors.lastname: Wiggins
authors.forename: Peter
authors.initials: P
authors.lastname: Tsvetkov
authors.forename: Mark
authors.initials: M
authors.lastname: Johnson
authors.forename: Claire L
authors.initials: CL
authors.lastname: Joyce
authors.forename: Christopher A
authors.initials: CA
authors.lastname: Lamb
authors.forename: Nia J
authors.initials: NJ
authors.lastname: Bryant
authors.forename: David
authors.initials: D
authors.lastname: Komander
authors.forename: Yosef
authors.initials: Y
authors.lastname: Shaul
authors.forename: Simon J
authors.initials: SJ
authors.lastname: Cook
mesh_terms: D000818:Animals; D051017:Apoptosis Regulatory Proteins / Q000235:genetics / Q000378:metabolism*; D000072224:Bcl-2-Like Protein 11; D002460:Cell Line; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism*; D051379:Mice; D018345:Mice, Knockout; D019950:Mitogen-Activated Protein Kinase 1 / Q000235:genetics / Q000378:metabolism; D048052:Mitogen-Activated Protein Kinase 3 / Q000235:genetics / Q000378:metabolism; D046988:Proteasome Endopeptidase Complex / Q000235:genetics / Q000378:metabolism*; D011518:Proto-Oncogene Proteins / Q000235:genetics / Q000378:metabolism*; D054875:Ubiquitination
chemical_list: D051017:Apoptosis Regulatory Proteins; D000072224:Bcl-2-Like Protein 11; C000606776:Bcl2l11 protein, mouse; D008565:Membrane Proteins; D011518:Proto-Oncogene Proteins; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; D046988:Proteasome Endopeptidase Complex